Ribonucleotide reductase is responsible for regulating a balanced and continued supply of deoxyribonucleotide precursors required for DNA replication. As an approach to understanding the structural and functional basis for regulation of ribonucleotide reduction, we have isolated and characterized a multienzyme complex containing ribonucleotide reductase proteins B1, B2 and thioredoxin. This form of ribonucleotide reductase is characterized as a highly active, and physically stable complex. Unlike the solubilized form of reductase which shows dilution inactivation, the specific activity in the membrane fraction is independent of protein concentration. The functional significance an the integrity of this membrane reductase complex is also suggested by copurification of several other enzymes involved in deoxynucleotide metabolism, including thymidylate synthetase and DNA polymerase I. Present studies will continue characterization of the various enzyme activities copurifying with the ribonucleotide reductase-membrane fraction together with studies on any functional relations that exist between these proteins. The role of thioredoxin as hydrogen donor has occupied our considerable attentions based on the findings that (i) this protein is phosphorylated on the thiols previously implicated in proton transfer; (ii) a substantial fraction of thioredoxin (activity and protein) appears physically associated with membranes. The ability of thioredoxin to serve both as hydrogen donor in vitro and the requirement for DNA synthesis (as an essential component to T7 replication) suggests that this protein may provide a key link in functioning of any multienzyme complex for deoxynucleotide biosynthesis. To establish this link we have pursued a better understanding of the subcellular localization of thioredoxin and the physiological role of phosphorylation.